Cytotoxic 2-benzylidene-6-(nitrobenzylidene)cyclohexanones which display substantially greater toxicity for neoplasms than non-malignant cells

Abstract

Various 2-benzylidene-6-(nitrobenzylidene)cyclohexanones were prepared as candidate cytotoxins in which the nitro group was located in the ortho, meta and para positions leading to series 1-3, respectively. The CC(50) values towards human HSC-2 and HSC-4 oral squamous cell carcinomas as well as human HL-60 promyelocytic leukemic cells are in the low micromolar range in general. On the other hand, most of the compounds afforded clear evidence of being far less toxic towards human HGF gingival fibroblasts, HPC pulp cells and HPLF periodontal ligament fibroblasts which are non-malignant cells. Selectivity index (SI) figures were generated which are the ratios of the average CC(50) values towards normal cells and the CC(50) figure towards a malignant cell line. Huge SI values were obtained for many of the compounds. In particular 1c, 2f, 3c and 3g which have average SI values of >76, >38, 124 and 341, respectively, are clearly lead molecules affording direction for amplification of this area of study. A lead compound 1c caused internucleosomal DNA fragmentation and activation of caspase-3 in HL-60 cells but not in HSC-2 carcinomas. In a short-term toxicity study, doses up to and including 300 mg/kg of the majority of the compounds prepared in this study did not cause any mortalities to mice. Some guidelines for development of these tumor-selective cytotoxins are presented.

Figure 1

(A) Design of the compounds in series 1–3. (B) Specific compounds prepared.

Figure 2

Structures of 5a–c.

Figure 3

Evaluation of 1c on the induction of internucleosomal DNA fragmentation in HSC-2 and HL-60 cells after 6 h of incubation. M is the molecular weight marker of DNA.

Figure 4

Effect of 1c on the activation of caspase-3 after 5 h incubation in HSC-2 and HL-60 cells. UV refers to ultraviolet radiation (6 J/m²/min) applied for 1 min followed by incubation of the cells for 3 h. The bar graphs represent the mean standard deviations (n = 3).